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ts mutations

David Michael Eisenmann eisenman at leland.Stanford.EDU
Thu Dec 5 13:09:29 EST 1996

Jorgensen at BIOSCIENCE.BIOLOGY.UTAH.EDU ("Jorgensen") writes:

>I have been wondering whether it is possible to engineer temperature
>sensitive mutations.  I made some of my own guesses about what kind of
>mutations would be temperature sensitive but when I talked to protein
>chemists, they had different ideas.  I was convinced that no one knew
>anything.  But then I found a paper in which the authors performed systematic
>mutagenesis of every amino acid in a bacterial protein and there were some
>generalities observed.  Basically, they saw that ts mutations typically
>substituted a charged residue for a leucine, isoleucine or valine contained
>in a hydrophobic stretch.  I wonder whether these general rules are true in
>C. elegans.  I would like to look at all the known ts mutations that have
>been sequenced in C elegans and let the empirical data guide us.  Phil
>Anderson lists the nucleotide changes for many mutations in his chapter in
>the protocols book but unfortunately this list doesn't identify the amino
>acid changes or whether they are temperature sensitive.  If any of you have
>sequenced ts mutations could you please tell me what the change was and
>whether it is in a hydrophillic or hydrophobic domain.

>Erik Jorgensen

Dear Erik,

	I have characterized a ts mutation in let-60 ras that causes a
ts Muv phenotype in vivo and a temperature-dependent GTPase activity in 
vitro.  It is a Leucine to Phenylalanine change at a residue buried in
a hydrophobic pocket of Ras.  The residues in this pocket are conserved
evolutionarily, and are believed to play structural roles (rather than
catalytic).  The change is L19F.  

Dave Eisenmann
eisenman at cmgm.stanford.edu

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